1887

Abstract

A stable plasmid DNA, pMWJEAT, was constructed by using full-length (JEV) cDNA isolated from the wild-type strain JEV AT31. Recombinant JEV was obtained by synthetic RNA transfection into Vero cells and designated rAT virus. JEV rAT exhibited similar large-plaque morphology and antigenicity to the parental AT31 strain. Mutant clone pMWJEAT-E138K, containing a single Glu-to-Lys mutation at aa 138 of the envelope (E) protein, was also constructed to analyse the mechanisms of viral attenuation arising from this mutation. Recombinant JEV rAT-E138K was also recovered and displayed a smaller-plaque morphology and lower neurovirulence and neuroinvasiveness than either AT31 virus or rAT virus. JEV rAT-E138K exhibited greater plaque formation than rAT virus in virus–cell interactions under acidic conditions. Heparin or heparinase III treatment inhibited binding to Vero cells more efficiently for JEV rAT-E138K than for rAT virus. Inhibition of virus–cell interactions by using wheatgerm agglutinin was more effective for JEV rAT than for rAT-E138K on Vero cells. About 20 % of macropinoendocytosis of JEV rAT for Vero cells was inhibited by cytochalasin D treatment, but no such inhibition occurred for rAT-E138K virus. Furthermore, JEV rAT was predominantly secreted from infected cells, whereas rAT-E138K was more likely to be retained in infected cells. This study demonstrates clearly that a single Glu-to-Lys mutation at aa 138 of the envelope protein affects multiple steps of the viral life cycle. These multiple changes may induce substantial attenuation of JEV.

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2005-08-01
2019-11-21
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References

  1. Aihara, S., Rao, C. M., Yu, Y. X., Lee, T., Watanabe, K., Komiya, T., Sumiyoshi, H., Hashimoto, H. & Nomoto, A. ( 1991; ). Identification of mutations that occurred on the genome of Japanese encephalitis virus during the attenuation process. Virus Genes 5, 95–109.[CrossRef]
    [Google Scholar]
  2. Allison, S. L., Schalich, J., Stiasny, K., Mandl, C. W., Kunz, C. & Heinz, F. X. ( 1995; ). Oligomeric rearrangement of tick-borne encephalitis virus envelope proteins induced by an acidic pH. J Virol 69, 695–700.
    [Google Scholar]
  3. Arroyo, J., Guirakhoo, F., Fenner, S., Zhang, Z.-X., Monath, T. P. & Chambers, T. J. ( 2001; ). Molecular basis for attenuation of neurovirulence of a yellow fever virus/Japanese encephalitis virus chimera vaccine (ChimeriVax-JE). J Virol 75, 934–942.[CrossRef]
    [Google Scholar]
  4. Chen, L.-K., Lin, Y.-L., Liao, C.-L., Lin, C.-G., Huang, Y.-L., Yeh, C.-T., Lai, S.-C., Jan, J.-T. & Chin, C. ( 1996; ). Generation and characterization of organ-tropism mutants of Japanese encephalitis virus in vivo and in vitro. Virology 223, 79–88.[CrossRef]
    [Google Scholar]
  5. Chen, Y., Maguire, T., Hileman, R. E., Fromm, J. R., Esko, J. D., Linhardt, R. J. & Marks, R. M. ( 1997; ). Dengue virus infectivity depends on envelope protein binding to target cell heparan sulfate. Nat Med 3, 866–871.[CrossRef]
    [Google Scholar]
  6. Gritsun, T. S. & Gould, E. A. ( 1998; ). Development and analysis of a tick-borne encephalitis virus infectious clone using a novel and rapid strategy. J Virol Methods 76, 109–120.[CrossRef]
    [Google Scholar]
  7. Gualano, R. C., Pryor, M. J., Cauchi, M. R., Wright, P. J. & Davidson, A. D. ( 1998; ). Identification of a major determinant of mouse neurovirulence of dengue virus type 2 using stably cloned genomic-length cDNA. J Gen Virol 79, 437–446.
    [Google Scholar]
  8. Hasegawa, H., Yoshida, M., Shiosaka, T., Fujita, S. & Kobayashi, Y. ( 1992; ). Mutations in the envelope protein of Japanese encephalitis virus affect entry into cultured cells and virulence in mice. Virology 191, 158–165.[CrossRef]
    [Google Scholar]
  9. Hayasaka, D., Gritsun, T. S., Yoshii, K. & 7 other authors ( 2004; ). Amino acid changes responsible for attenuation of virus neurovirulence in an infectious cDNA clone of the Oshima strain of Tick-borne encephalitis virus. J Gen Virol 85, 1007–1018.[CrossRef]
    [Google Scholar]
  10. Heinz, F. X. ( 1986; ). Epitope mapping of flavivirus glycoproteins. Adv Virus Res 31, 103–168.
    [Google Scholar]
  11. Hung, S.-L., Lee, P.-L., Chen, H.-W., Chen, L.-K., Kao, C.-L. & King, C.-C. ( 1999; ). Analysis of the steps involved in dengue virus entry into host cells. Virology 257, 156–167.[CrossRef]
    [Google Scholar]
  12. Hung, J.-J., Hsieh, M.-T., Young, M.-J., Kao, C.-L., King, C.-C. & Chang, W. ( 2004; ). An external loop region of domain III of dengue virus type 2 envelope protein is involved in serotype-specific binding to mosquito but not mammalian cells. J Virol 78, 378–388.[CrossRef]
    [Google Scholar]
  13. Hurrelbrink, R. J. & McMinn, P. C. ( 2001; ). Attenuation of Murray Valley encephalitis virus by site-directed mutagenesis of the hinge and putative receptor-binding regions of the envelope protein. J Virol 75, 7692–7702.[CrossRef]
    [Google Scholar]
  14. Hurrelbrink, R. J., Nestorowicz, A. & McMinn, P. C. ( 1999; ). Characterization of infectious Murray Valley encephalitis virus derived from a stably cloned genome-length cDNA. J Gen Virol 80, 3115–3125.
    [Google Scholar]
  15. Kato, T., Miyamoto, M., Furusaka, A., Date, T., Yasui, K., Kato, J., Matsushima, S., Komatsu, T. & Wakita, T. ( 2003a; ). Processing of hepatitis C virus core protein is regulated by its C-terminal sequence. J Med Virol 69, 357–366.[CrossRef]
    [Google Scholar]
  16. Kato, T., Date, T., Miyamoto, M., Furusaka, A., Tokushige, K., Mizokami, M. & Wakita, T. ( 2003b; ). Efficient replication of the genotype 2a hepatitis C virus subgenomic replicon. Gastroenterology 125, 1808–1817.[CrossRef]
    [Google Scholar]
  17. Khromykh, A. A. & Westaway, E. G. ( 1994; ). Completion of Kunjin virus RNA sequence and recovery of an infectious RNA transcribed from stably cloned full-length cDNA. J Virol 68, 4580–4588.
    [Google Scholar]
  18. Kimura-Kuroda, J. & Yasui, K. ( 1983; ). Topographical analysis of antigenic determinants on envelope glycoprotein V3 (E) of Japanese encephalitis virus, using monoclonal antibodies. J Virol 45, 124–132.
    [Google Scholar]
  19. Kimura-Kuroda, J., Ichikawa, M., Ogata, A., Nagashima, K. & Yasui, K. ( 1993; ). Specific tropism of Japanese encephalitis virus for developing neurons in primary rat brain culture. Arch Virol 130, 477–484.[CrossRef]
    [Google Scholar]
  20. Kinney, R. M., Butrapet, S., Chang, G.-J. J., Tsuchiya, K. R., Roehrig, J. T., Bhamarapravati, N. & Gubler, D. J. ( 1997; ). Construction of infectious cDNA clones for dengue 2 virus: strain 16681 and its attenuated vaccine derivative, strain PDK-53. Virology 230, 300–308.[CrossRef]
    [Google Scholar]
  21. Kolaskar, A. S. & Kulkarni-Kale, U. ( 1999; ). Prediction of three-dimensional structure and mapping of conformational epitopes of envelope glycoprotein of Japanese encephalitis virus. Virology 261, 31–42.[CrossRef]
    [Google Scholar]
  22. Kuhn, R. J., Zhang, W., Rossmann, M. G. & 9 other authors ( 2002; ). Structure of dengue virus: implications for flavivirus organization, maturation, and fusion. Cell 108, 717–725.[CrossRef]
    [Google Scholar]
  23. Lai, C.-J., Zhao, B., Hori, H. & Bray, M. ( 1991; ). Infectious RNA transcribed from stably cloned full-length cDNA of dengue type 4 virus. Proc Natl Acad Sci U S A 88, 5139–5143.[CrossRef]
    [Google Scholar]
  24. Lee, E. & Lobigs, M. ( 2002; ). Mechanism of virulence attenuation of glycosaminoglycan-binding variants of Japanese encephalitis virus and Murray Valley encephalitis virus. J Virol 76, 4901–4911.[CrossRef]
    [Google Scholar]
  25. Lee, E., Hall, R. A. & Lobigs, M. ( 2004; ). Common E protein determinants for attenuation of glycosaminoglycan-binding variants of Japanese encephalitis and West Nile viruses. J Virol 78, 8271–8280.[CrossRef]
    [Google Scholar]
  26. Lescar, J., Roussel, A., Wien, M. W., Navaza, J., Fuller, S. D., Wengler, G., Wengler, G. & Rey, F. A. ( 2001; ). The fusion glycoprotein shell of Semliki Forest virus: an icosahedral assembly primed for fusogenic activation at endosomal pH. Cell 105, 137–148.[CrossRef]
    [Google Scholar]
  27. Liu, H., Chiou, S.-S. & Chen, W.-J. ( 2004; ). Differential binding efficiency between the envelope protein of Japanese encephalitis virus variants and heparan sulfate on the cell surface. J Med Virol 72, 618–624.[CrossRef]
    [Google Scholar]
  28. Mandl, C. W., Guirakhoo, F., Holzmann, H., Heinz, F. X. & Kunz, C. ( 1989; ). Antigenic structure of the flavivirus envelope protein E at the molecular level, using tick-borne encephalitis virus as a model. J Virol 63, 564–571.
    [Google Scholar]
  29. Mandl, C. W., Kroschewski, H., Allison, S. L., Kofler, R., Holzmann, H., Meixner, T. & Heinz, F. X. ( 2001; ). Adaptation of tick-borne encephalitis virus to BHK-21 cells results in the formation of multiple heparan sulfate binding sites in the envelope protein and attenuation in vivo. J Virol 75, 5627–5637.[CrossRef]
    [Google Scholar]
  30. Modis, Y., Ogata, S., Clements, D. & Harrison, S. C. ( 2004; ). Structure of the dengue virus envelope protein after membrane fusion. Nature 427, 313–319.[CrossRef]
    [Google Scholar]
  31. Monath, T. P. & Heinz, F. X. ( 1996; ). Flaviviruses. In Fields Virology, 3rd edn, pp. 961–1034. Edited by B. N. Fields, D. M. Knipe & P. M. Howley. Philadelphia, PA: Lippincott-Raven.
  32. Monath, T. P., Arroyo, J., Levenbook, I., Zhang, Z.-X., Catalan, J., Draper, K. & Guirakhoo, F. ( 2002; ). Single mutation in the flavivirus envelope protein hinge region increases neurovirulence for mice and monkeys but decreases viscerotropism for monkeys: relevance to development and safety testing of live, attenuated vaccines. J Virol 76, 1932–1943.[CrossRef]
    [Google Scholar]
  33. Ni, H. & Barrett, A. D. T. ( 1998; ). Attenuation of Japanese encephalitis virus by selection of its mouse brain membrane receptor preparation escape variants. Virology 241, 30–36.[CrossRef]
    [Google Scholar]
  34. Nitayaphan, S., Grant, J. A., Chang, G.-J. J. & Trent, D. W. ( 1990; ). Nucleotide sequence of the virulent SA-14 strain of Japanese encephalitis virus and its attenuated vaccine derivative, SA-14-14-2. Virology 177, 541–552.[CrossRef]
    [Google Scholar]
  35. Polo, S., Ketner, G., Levis, R. & Falgout, B. ( 1997; ). Infectious RNA transcripts from full-length dengue virus type 2 cDNA clones made in yeast. J Virol 71, 5366–5374.
    [Google Scholar]
  36. Reed, L. J. & Muench, H. ( 1938; ). A simple method of estimating fifty percent endpoints. Am J Hyg 27, 493–497.
    [Google Scholar]
  37. Rey, F. A., Heinz, F. X., Mandl, C., Kunz, C. & Harrison, S. C. ( 1995; ). The envelope glycoprotein from tick-borne encephalitis virus at 2 Å resolution. Nature 375, 291–298.[CrossRef]
    [Google Scholar]
  38. Rice, C. M., Grakoui, A., Galler, R. & Chambers, T. J. ( 1989; ). Transcription of infectious yellow fever virus RNA from full-length cDNA templates produced by in vitro ligation. New Biol 1, 285–296.
    [Google Scholar]
  39. Shi, P.-Y., Tilgner, M., Lo, M. K., Kent, K. A. & Bernard, K. A. ( 2002; ). Infectious cDNA clone of the epidemic West Nile virus from New York City. J Virol 76, 5847–5856.[CrossRef]
    [Google Scholar]
  40. Stiasny, K., Allison, S. L., Marchler-Bauer, A., Kunz, C. & Heinz, F. X. ( 1996; ). Structural requirements for low-pH-induced rearrangements in the envelope glycoprotein of tick-borne encephalitis virus. J Virol 70, 8142–8147.
    [Google Scholar]
  41. Stiasny, K., Allison, S. L., Schalich, J. & Heinz, F. X. ( 2002; ). Membrane interactions of the tick-borne encephalitis virus fusion protein E at low pH. J Virol 76, 3784–3790.[CrossRef]
    [Google Scholar]
  42. Sumiyoshi, H., Mori, C., Fuke, I., Morita, K., Kuhara, S., Kondou, J., Kikuchi, Y., Nagamatu, H. & Igarashi, A. ( 1987; ). Complete nucleotide sequence of the Japanese encephalitis virus genome RNA. Virology 161, 497–510.[CrossRef]
    [Google Scholar]
  43. Sumiyoshi, H., Hoke, C. H. & Trent, D. W. ( 1992; ). Infectious Japanese encephalitis virus RNA can be synthesized from in vitro-ligated cDNA templates. J Virol 66, 5425–5431.
    [Google Scholar]
  44. Sumiyoshi, H., Tignor, G. H. & Shope, R. E. ( 1995; ). Characterization of a highly attenuated Japanese encephalitis virus generated from molecularly cloned cDNA. J Infect Dis 171, 1144–1151.[CrossRef]
    [Google Scholar]
  45. Wang, T., Town, T., Alexopoulou, L., Anderson, J. F., Fikrig, E. & Flavell, R. A. ( 2004; ). Toll-like receptor 3 mediates West Nile virus entry into the brain causing lethal encephalitis. Nat Med 10, 1366–1373.[CrossRef]
    [Google Scholar]
  46. Whitehead, S. S., Falgout, B., Hanley, K. A., Blaney, J. E., Jr, Markoff, L. & Murphy, B. R. ( 2003; ). A live, attenuated dengue virus type 1 vaccine candidate with a 30-nucleotide deletion in the 3′ untranslated region is highly attenuated and immunogenic in monkeys. J Virol 77, 1653–1657.[CrossRef]
    [Google Scholar]
  47. Wilcox, C. L., Smith, R. L., Freed, C. R. & Johnson, E. M., Jr ( 1990; ). Nerve growth factor-dependence of herpes simplex virus latency in peripheral sympathetic and sensory neurons in vitro. J Neurosci 10, 1268–1275.
    [Google Scholar]
  48. Yamshchikov, V., Mishin, V. & Cominelli, F. ( 2001; ). A new strategy in design of (+)RNA virus infectious clones enabling their stable propagation in E. coli. Virology 281, 272–280.[CrossRef]
    [Google Scholar]
  49. Yasui, K. ( 2002; ). Neuropathogenesis of Japanese encephalitis virus. J Neurovirol 8 (Suppl. 2), 112–114.[CrossRef]
    [Google Scholar]
  50. Yoshii, K., Konno, A., Goto, A. & 7 other authors ( 2004; ). Single point mutation in tick-borne encephalitis virus prM protein induces a reduction of virus particle secretion. J Gen Virol 85, 3049–3058.[CrossRef]
    [Google Scholar]
  51. Yu, Y. X., Wu, P. F., Ao, J., Liu, L. H. & Li, H. M. ( 1981; ). Selection of a better immunogenic and highly attenuated live vaccine virus strain of JE. I. Some biological characteristics of SA14-14-2 mutant. Chin J Microbiol Immunol 1, 77–84.
    [Google Scholar]
  52. Yun, S.-I., Kim, S.-Y., Rice, C. M. & Lee, Y.-M. ( 2003; ). Development and application of a reverse genetics system for Japanese encephalitis virus. J Virol 77, 6450–6465.[CrossRef]
    [Google Scholar]
  53. Zhang, W., Chipman, P. R., Corver, J. & 7 other authors ( 2003a; ). Visualization of membrane protein domains by cryo-electron microscopy of dengue virus. Nat Struct Biol 10, 907–912.[CrossRef]
    [Google Scholar]
  54. Zhang, Y., Corver, J., Chipman, P. R. & 7 other authors ( 2003b; ). Structures of immature flavivirus particles. EMBO J 22, 2604–2613.[CrossRef]
    [Google Scholar]
  55. Zhao, Z., Wakita, T. & Yasui, K. ( 2003; ). Inoculation of plasmids encoding Japanese encephalitis virus PrM-E proteins with colloidal gold elicits a protective immune response in BALB/c mice. J Virol 77, 4248–4260.[CrossRef]
    [Google Scholar]
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vol. , part 8, pp. 2209 – 2220

Construction of the full-length cDNA clone pMWJEAT

E protein sequences of nine flaviviruses aligned to the JEV AT31 E protein sequence in the region surrounding residue 138

List of primers used in this study

Binding efficiency between pairs of mosquito-borne flavivirus variants containing the Glu-to-Lys mutation at residue E-138 and heparan sulphate

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